PROJECT SUMMARY/ABSTRACT The development of type 1 diabetes (T1D) relies on interactions between genes imparting disease susceptibility and environmental factors thought to contribute to aberrant immune activation and signaling leading to a breakdown in tolerance. These complex interactions adversely affect the development and function of the immune cells, which mediate the autoimmune destruction of insulin-producing pancreatic ?- cells. In addition to the human leukocyte antigen (HLA) region, over 50 genetic loci have been identified that confer varying degrees of risk for T1D. However, the specific mechanisms by which these variants alter the development and signaling events within in the immune system remain poorly characterized. Our studies are designed to address these knowledge gaps through investigation of peripheral blood and tissues derived from a diverse cohort of study subjects (Core B). Through these investigations, we hope to gain a better understanding for how genetic risk variants influence responses to environmental stimuli (e.g., IFN1; Project 1) that lead to a large degree of heterogeneity observed in the natural history and clinical manifestations of the disease. We will test the hypothesis that a combination of susceptibility gene variants and aberrant extrinsic growth, survival, and differentiation factors lead to a loss of T cell tolerance in the context of T1D. We propose to conduct a series of Aims to: 1) identify the T1D-associated cellular and phenotypic signatures in a cross-sectional cohort of subjects with T1D, their relatives at varying degrees of risk for the disease, and controls using extensive human immunophenotyping (HIP) by flow cytometry together with genome-wide genotyping of >974K single nucleotide polymorphisms (SNPs) and sophisticated bioinformatics analyses (with Project 3); 2) determine the phenotypic and functional impact of genetic risk variants and age-associated growth factors on regulatory and effector T cell function, and 3) create T cell receptor (TCR) ?avatars? with known reactivities and isogenic cellular systems to develop a ?disease in a dish? model for assessing antigen-specific T cell function. These latter studies will employ novel tools including lentiviral expression systems, directed gene editing of human T cells, and the utilization of unique clinical resources to address the challenge of epistatic gene interactions in humans with T1D. In sum, data from these Aims are expected to provide essential information about the complex interactions between genetic risk, immune signaling events, and immune cell development that impact autoimmune disease pathogenesis. The development and utilization of these innovative platforms will expedite our ability to identify and test novel therapeutic interventions to halt the autoimmune destruction of ?-cells in T1D.